U.S. patent number 5,766,957 [Application Number 08/716,213] was granted by the patent office on 1998-06-16 for spectrophotometric techniques.
This patent grant is currently assigned to Applied Research Systems Ars Holding, N.V.. Invention is credited to Alan Derek Cookson, Grenville Arthur Robinson.
United States Patent |
5,766,957 |
Robinson , et al. |
June 16, 1998 |
Spectrophotometric techniques
Abstract
The use as a cuvette in spectrophotometric assays of a
sample-collecting device is disclosed, the said device possessing a
cavity or cavities each having a dimension small enough to enable
sample liquid to be drawn into the cavity by capillary action,
wherein a surface of the cavity is a surface of a transparent solid
plate having a substantially rectangular cross section and forming
a wall of the cavity, the opposite surface of the cavity being an
additional structure having a substantially rectangular cross
section and forming a wall of the cavity, and wherein a portion of
said plate carries a reflective coating and a portion of said
additional structure is reflective.
Inventors: |
Robinson; Grenville Arthur
(Ealing, GB), Cookson; Alan Derek (Boston, MA) |
Assignee: |
Applied Research Systems Ars
Holding, N.V. (Curacao, NL)
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Family
ID: |
10751683 |
Appl.
No.: |
08/716,213 |
Filed: |
November 22, 1996 |
PCT
Filed: |
March 09, 1995 |
PCT No.: |
PCT/GB95/00507 |
371
Date: |
November 22, 1996 |
102(e)
Date: |
November 22, 1996 |
PCT
Pub. No.: |
WO95/24632 |
PCT
Pub. Date: |
September 14, 1995 |
Foreign Application Priority Data
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Mar 10, 1994 [GB] |
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9404749 |
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Current U.S.
Class: |
436/165; 356/246;
356/440; 422/82.09; 422/82.11 |
Current CPC
Class: |
B01L
3/502715 (20130101); G01N 21/0303 (20130101); B01L
2300/0822 (20130101); B01L 2300/168 (20130101); B01L
2400/0406 (20130101) |
Current International
Class: |
B01L
3/00 (20060101); G01N 21/03 (20060101); G01N
027/26 () |
Field of
Search: |
;422/82.09,82.11
;436/164,165,180,171 ;356/246,440 |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
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0171148 |
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Feb 1986 |
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EP |
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9014590 |
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Nov 1990 |
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WO |
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Other References
Analytical Chemistry, vol. 55, No. 6, 1983 Easton Pennsylvania,
USA, pp. 951-955, Wei Lai et al. "Determination of phosphorus in
natural waters by long capillaryC cell absorption
spectrometry"..
|
Primary Examiner: Snay; Jeffrey
Attorney, Agent or Firm: Ostrolenk, Faber, Gerb &
Soffen, LLP
Claims
We claim:
1. A method of spectrophotometric assay of a species in a liquid
sample using a sample-collecting device comprising a cavity or
cavities each having a dimension small enough to have a sample
liquid drawn into the cavity by capillary action, wherein a surface
of the cavity is a surface of a transparent solid plate having a
substantially rectangular cross-section and forming a wall of the
cavity, the opposite surface of the cavity is an additional
structure having a substantially rectangular cross-section and
forming a wall of the cavity, and wherein a portion of said plate
carries a reflective coating and a portion of said additional
structure is reflective, said method comprising
(a) filling said device with the sample liquid;
(b) irradiating said device with light from a suitable light source
such that the radiation is totally internally reflected within said
device; and
(c) monitoring the radiation emerging from said device in order to
determine whether and optionally the extent to which the species is
present in the sample.
2. A method as claimed in claim 1 wherein the additional structure
of the device is also a transparent plate having a portion of which
carries a reflective coating.
3. A method as claimed in claim 1 wherein a reflective coating is
on an external surface of said plate.
4. A cuvette as claimed in claim 1, wherein a reflective coating is
on each external surface of said plate.
5. A method as claimed in claim 1 wherein the device has a length
of between about 20 mm and 60 mm, a width of between about 5 mm and
20 mm and a gap between the transparent plate and the additional
structure of about 0.1 and 1 mm.
6. A method as claimed in claim 5, wherein the length is about 40
mm, the width is about 10 mm and the gap is about 0.1 to 0.5
mm.
7. A method as claimed in claim 6, wherein the additional structure
of the device is a second transparent plate having a portion of
which which carries a reflecting coating.
8. A method as claimed in claim 7, wherein a reflecting coating is
on each external surface of said second plate.
Description
BACKGROUND OF THE INVENTION
The present invention relates to spectrophotometric techniques
using particular devices.
In EP-A-0171148, devices are disclosed for use in chemical
(especially biochemical or clinical) test procedures, particularly
for use in specific binding assay procedures. The devices typically
possess a cavity into which sample liquid is drawn. One surface of
the cavity is a surface of a transparent solid plate which acts as
a light transmissive waveguide; for use in specific binding assays,
this surface carries an immobilised reagent appropriate to the
assay to be carried out in the device. The plate has an edge which
is substantially optically smooth and transverse to the plane of
the plate and in use in an assay procedure the device can transmit,
for example, fluorescence from an adsorbed fluorescent material
along the plate acting as a waveguide, transfer of useful light
across the boundary of the plate occurring by the evanescent wave
located very close to the interface.
We have now found that devices similar to those in EP-A-0171148 may
be used as a cuvette in spectrophotometric assays. In the normal
mode of use of the devices as described in EP-A-0171148, there is
only a thin film of sample (typically of the order of 0.1 mm)
present in the device; passing radiation through a path length of
this dimension is not suitable for reliable spectrophotometric
measurement. We have now found, however, that totally internally
reflecting radiation within a modified form of the device allows
the radiation to pass through the sample many times and therefore
enables a suitable path length (for example, of up to 1 cm) to be
built up. Such a path length is then suitable for reliable
spectrophotometric measurement.
Devices for analysing substances and which employ multiple internal
reflection are already known. In U.S. Pat. No. 5,273,633 (Tiansong
Wang), a device for capillary zone electrophoresis is disclosed
which comprises a long transparent capillary tube with a reflecting
surface surrounding the outer surface of the capillary, the
reflecting surface having an incident and exit window for the
electromagnetic radiation. Similarly, in Analytical Chem., vol 55,
no. 6, 1983, pp 951-955 (Wei Lei et al), a long capillary cell, in
which in use multiple internal reflection of electromagnetic
radiation occurs, is employed for the colorimetric determination of
phosphorus. U.S. Pat. No. 3,431,424 (Henry W. Allen) discloses a
complex optical fluid sampling device which in use also empoys
multiply reflected radiation.
These prior art devices are all based on a circular geometry and
this presents certain disadvantages. As specifically acknowledged
in U.S. Pat. No. 5,273,633, the fact that radial reflection occurs
places constraints on the size and orientation of the incident
window and also, since the exiting radiation will be distributed
around the entire circumference of the capillary, on the preferred
shape of the exit window and the detection apparatus. Furthermore,
these prior art devices are relatively large (of the order of
50-100 cm) and this makes them impractical for use in routine
analytical work. The use according to the present invention of a
device as described above (the modified form of the device
disclosed in EP-A-0171148) overcomes these disadvantages.
DESCRIPTION OF THE INVENTION
Thus, according to one aspect of the present invention, we provide
the use of a sample-collecting device as a cuvette in
spectrophotometric assays, the device possessing a cavity or
cavities each having a dimension small enough to enable sample
liquid to be drawn into the cavity by capillary action, wherein a
surface of the cavity is a surface of a transparent solid plate
having a substantially rectangular cross-section and forming a wall
of the cavity, the opposite surface of the cavity being an
additional structure having a substantially rectangular
cross-section and forming a wall of the cavity, and wherein a
portion of said plate carries a reflective coating and a portion of
said additional structure is reflective.
According to a further aspect of the present invention there is
provided a method of spectrophotometric assay of a species in a
liquid sample using a device as defined above which comprises
(a) filling said device with the sample liquid;
(b) irradiating said device with light from a suitable light source
such that the radiation is totally internally reflected within said
device; and
(c) monitoring the radiation emerging from said device by
conventional methods in order to determine whether and/or the
extent to which the species is present in the sample.
Preferably the additional structure of the device is also a
transparent plate a portion of which carries a reflective coating.
However, this is not a necessary requirement and it is suitable
merely to use a structure part of which is fabricated of an
appropriate reflective material; in such an embodiment the material
concerned is preferably inert with regard to the sample liquid to
be contained within the cavity.
The reflective coating carried on the or each transparent plate is
preferably carried on the external surface i.e. the surface remote
from the cavity. However, the coating on the or each transparent
plate may instead be carried on the internal surface i.e. the
surface opposite the external surface (being that which forms part
of the capillary cavity wall). Alternatively, where two transparent
plates are present, one may carry the coating on the external
surface whereas the other may carry the coating on the internal
surface.
The reflective portions of the transparent plate and of the
additional structure facilitate total internal reflection of
radiation within the device in use in order to build up a suitable
path length of the radiation through the sample.
The choice of where the reflective coating is carried on the or
each transparent plate is made in the light of this result to be
achieved.
A suitable coating for the transparent plate(s) is a metal coating
as conventionally used in the preparation of coated glass, e.g. an
aluminium coating.
The devices for use according to the present invention are
conveniently manufactured, enable convenient sample collection by
virtue of their capillarity and are especially suitable for
handling and accurately metering small liquid samples, the volume
of the cavity containing the sample being of the order of 0.1 ml in
typical embodiments.
The devices for use according to the present invention may be
constructed by similar methods to those described in detail for the
devices of EP-A-0171148, but with the additional step of applying a
reflective coating to the appropriate surface of the transparent
plate(s).
Thus, typically a process for manufacturing sample-collecting
devices as described hereinbefore would comprise the steps of
(a) applying a coating of reflective material to a portion of a
surface of a transparent sheet material which is to provide part of
a multiplicity of the devices;
(b) either prior to or subsequent to step (a), attaching to said
sheet material in parallel, spaced relation thereto an additional
structure which together with said sheet material provides for each
device of the multiplicity of devices a cavity of capillary
dimension for collecting and retaining by capillarity a volume of
sample liquid, a portion of the additional structure being
reflective; and
(c) separating the assembled laminate into portions each providing
one or a plurality of the sample-collecting devices.
Where the additional structure is a transparent sheet material, a
portion of which carries a coating of reflective material, an
additional step would comprise the application of this coating.
The orientation of the attachment in (b) will determine on which
surfaces (external or internal) of the cavity the coating(s) of
reflective material is(are) carried.
The transparent plate(s) of the device for use according to the
present invention may be fabricated from a variety of materials
which can be transparent to infrared, visible and/or ultraviolet
light depending on the spectrophotometric method to be carried out
using the device. Suitable materials are, for example, glass e.g.
sheet soda glass about 1 mm thick, silicas and plastics sheet
material e.g. acrylic plastics material as well as materials
conventionally used in the fabrication of spectroscopic cells such
as quartz, NaCl and CaF.sub.2. Where part of the additional
structure is simply fabricated from a reflective material, this
may, for example, be a chemically inert reflective metal such as
gold or platinum.
A number of modifications of the devices as disclosed in detail in
EP-A-0171148 are applicable to the device for use according to the
present invention including for example the use of a barrier for
selective takeup of sample liquid into the cavity, the use of a
collection surface of the device onto which a drop of sample liquid
can be placed and the provision of a fixed or releasable holder for
the device. The present invention extends to the use of devices as
described above incorporating such features.
The sample-collecting device as defined above would be used in
conventional spectrophotometric detection methods and would find
particular utility in the detection and determination of chemical
moieties in biological samples. For example the devices are able to
draw up a very small sample of blood e.g. from a finger spot and
thus analysis of such a sample for routine moieties including
haemoglobin, glucose and urea is envisaged.
In certain test methods, ancillary reagents may be needed and these
can be dosed separately, or they can be carried in dry releasable
form on a part of the device to be contacted in use by sample
liquid e.g. a surface of the capillary cavity or a surface of a
filter, if present. Such techniques are illustrated in
EP-A-0171148.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 shows a diagrammatic section through a device according to
one embodiment of the invention.
FIG. 2 shows a diagrammatic plan of the device of FIG. 1, and
includes a line I--I to show the line of section of FIG. 1.
FIGS. 3 and 4 show schematically the use of the device according to
the invention.
The device illustrated in FIGS. 1 and 2 comprises a transparent
plate 1 and a transparent plate 2 fixed together in parallel
opposed and spaced relation, preferably less than 1 mm apart, by
bonding tracks 3 of suitable (e.g. epoxy) adhesive to form a
capillary cell cavity 4, open at both ends, which communicates with
the outside though a first discontinuity in the bonding 3 arranged
to form a cell aperture at side 5 of plate 1. Another discontinuity
is present at the other end of bonding 3, to leave another
aperture, to allow exit of air when a sample liquid is loaded into
the cell. Plate 2 has a portion 6 extending away from the aperture
which acts as a platform or threshold or lip onto which a drop of
sample liquid can be applied, so that this liquid can be made to
fill the capillary cell cavity 4 by capillary flow. Cavity 4
attracts and contains a definite and adequately reproducible volume
of liquid when loaded in this way. On a portion of each external
surface of both plate 1 and plate 2 are respectively carried
reflective coatings 7 and 8.
In the manufacture of such a device, the coating of the outside
surfaces of the plates is carried out by conventional methods e.g.
by the use of a vacuum and evaporation of a suitable metal such as
silver or aluminium. The sealing is preferably achieved by screen
printing onto one plate lines of epoxy resin which suitably
comprise solid particles to ensure the desired spacing (e.g.
substantially monodisperse ballotini). The two sheets are then
brought together, subjected to vacuum and the adhesive cured by
ultraviolet illumination. The plates are then scribed and broken
into individual cell units.
Devices for use according to the present invention are preferably
fabricated with a gap of between 0.1 mm and 1 mm, preferably to 0.5
mm, more preferably about 0.2 mm. The transparent plate(s)
preferably have a thickness of between 0.5 and 1.5 mm, more
preferably about 1.0 mm. The devices are also preferably fabricated
so that in use in spectrophotometric assays the path length of the
radiation through the sample is between 0.1 and 2.0 cm, more
preferably about 1.0 cm. The devices are preferably of a length
between 20 mm and 60 mm, more preferably about 40 mm, and have a
width of between 5 mm and 20 mm, more preferably about 10 mm.
Thus, according to a further aspect of the present invention we
provide a sample-collecting device as defined above in which the
device has a length of between about 20 mm and 60 mm, a width of
between about 5 mm and 20 mm and a gap between the transparent
plate and the additional structure of about 0.1 mm to 1 mm.
FIG. 3 schematically illustrates the use of a device according to
the present invention in spectrophotometric measurement, the
incoming radiation being internally reflected in the device and
exiting after a suitable path length has been built up.
The path length through the sample would be controlled by the
length of the device, the thickness of the plates, the positioning
of the reflective metal layers on the surfaces of the device and
the angle of incidence of the radiation. Placing one or both of the
coatings on the internal surface of the plate(s) as opposed to the
external surface would also affect the overall path length, since
it would alter the skip distance for each bounce of radiation
within the device.
For example, for a device as illustrated in FIG. 3 where the width
of each transparent plate is 1.1 mm and the angle of incidence of
the radiation is 20.degree., each bounce of the light would result
in a skip distance of approximately 1.75 mm (4.8.times.tan
20.degree.) down the device with the light path through the sample
being approximately 0.42 mm. Thus a device of this type which is at
least 35 mm long would have a path length through the sample of at
least 0.8 cm. In practice the device would be slightly longer, for
example in the region of 40 mm, to allow for ease of handling.
Assuming that the internal width of the device is 10 mm (between
the bonding tracks) then the sample volume would be approximately
80 .mu.l.
As an alternative configuration it is possible to allow for a
sample and reference beam within the device, as illustrated
schematically in FIG. 4. Ideally one light source which is split
into two and chopped would be used to illuminate both sides of the
device. The device would be formed with two separate cavities, one
cavity containing the sample of interest and through which the
sample beam travels, the other cavity containing the reference
solution (essentially the `solvent` of the sample solution) through
which the reference beam passes. As an alternative, two distinct
and separate devices could be used akin to a conventional double
beam spectrophotometric technique. In either case, the use of a
single detector is beneficial to avoid the problems of matching
more than one detector.
* * * * *